One of my long-term research interests has been to understand the process of how cytokine signaling correlates with biological functions and the mechanisms of cytokine dysregulation in human diseases. The overall goal of this program is to identify signaling molecules and genes that may determine cytokine specificity and redundancy. In the past granting period, we have focused on studying the signal transduction mechanisms mediated by interleukin (IL)-9, a cytokine first cloned by the PI in 1988. We have compared the signaling events mediated by IL-9, which is an IL-2 receptor gamma chain (IL-2Rgamma,) superfamily cytokine, and other cytokines (such as IL-4) within the same superfamily. We have identified several signaling molecules which may determine cytokine specificity (such as STATs, IRSs, Tip60, 14-3-3) and redundancy (such as Cited2) for lL-2Rgamma superfamily cytokines. In the next granting period, we would like to further pursue the molecules, which determine specificity between IL-4 and IL-9 based on several novel findings uncovered during the past granting period. We have shown that (1) Tip60 (Tat-interacting protein, 60 kDa) can be tyrosine phosphorylated by Jak1 and Jak3 following IL-9, but not IL-4, stimulation, (2) Tip60 is rapidly degraded in an ubiquitination-dependent manner, and tyrosine phosphorylation of Tip60 is required for the stabilization of the Tip60 protein, (3) Tip60 complexes with endogenous Stat3, and regulates nuclear translocation of STAT3 induced by IL-9, (4) Tip60 acts as a co-repressor for STAT3 to regulate gene expression by the recruitment of a deacetylase, HDAC7, (5) STAT3 is acetylated following IL-9 stimulation and the acetylation can be abrogated by Tip60 HAT- mutant and (6) Nmi and importin-alpha7, two proteins shown to be involved in the signaling of IFN-gamma, and IL-2, are newly identified Tip60 interacting proteins. Based on our novel findings and those of others, we hypothesize that (1) Tip60/IL-9Ralpha interaction and STAT3 acetylation are important for STAT3-mediated functions and (2) characterization of Tip60 interacting proteins such as Nmi and importin-alpha7 may uncover novel mechanisms of nuclear transport and activation/inactivation of STATs. To test these hypotheses, we will (1) Study the mechanism and biological significance of Tip60/IL-9Ralpha interaction and STAT3 acetylation by analyzing the effects of acetylated STAT3 on nuclear translocation, DNA-binding, transcriptional activation and biological functions mediated by STAT3 and (II) Study Tip60 and its interacting proteins in STAT activation by analyzing the effects of Nmi and importin-alpha7 on nuclear translocation, DNA-binding, transcription activation and biological functions mediated by STAT3/Tip60. This proposal combines several important research areas of current interests including cytokine specificity/redundancy, nuclear import/export, protein phosphorylation/ ubiquitination/ acetylation/ deacetylation, transcriptional activation/repression and cytokine-mediated cellular functions. It is anticipated the accomplishment of the proposal will further our understanding in the control mechanisms of cytokine signal transduction which may lead to new modalities of treatment for cancer and leukemia/lymphoma. [unreadable] [unreadable] [unreadable]